Li Youfen, Park Jeong-Soon, Deng Jian-Hong, Bai Yidong
Department of Cellular and Structural Biology, University of Texas Health Sciences Center at San Antonio, 7703 Floyd Curl, San Antonio, Texas 78229, USA.
J Bioenerg Biomembr. 2006 Dec;38(5-6):283-91. doi: 10.1007/s10863-006-9052-z.
Cytochrome c oxidase or complex IV, catalyzes the final step in mitochondrial electron transfer chain, and is regarded as one of the major regulation sites for oxidative phosphorylation. This enzyme is controlled by both nuclear and mitochondrial genomes. Among its 13 subunits, three are encoded by mitochondrial DNA and ten by nuclear DNA. In this work, an RNA interference approach was taken which led to the generation of mouse A9 cell derivatives with suppressed expression of nuclear-encoded subunit IV (COX IV) of this complex. The amounts of this subunit are decrease by 86% to 94% of normal level. A detail biosynthetic and functional analysis of several cell lines with suppressed COX IV expression revealed a loss of assembly of cytochrome c oxidase complex and, correspondingly, a reduction in cytochrome c oxidase-dependent respiration and total respiration. Furthermore, dysfunctional cytochrome c oxidase in the cells leads to a compromised mitochondrial membrane potential, a decreased ATP level, and failure to grow in galactose medium. Interestingly, suppression of COX IV expression also sensitizes the cells to apoptosis. These observations provide the evidence of the essential role of the COX IV subunit for a functional cytochrome c oxidase complex and also demonstrate a tight control of cytochrome c oxidase over oxidative phosphorylation. Finally, our results further shed some insights into the pathogenic mechanism of the diseases caused by dysfunctional cytochrome c oxidase complex.
细胞色素c氧化酶或复合体IV催化线粒体电子传递链的最后一步，被视为氧化磷酸化的主要调控位点之一。这种酶受核基因组和线粒体基因组的共同控制。在其13个亚基中，3个由线粒体DNA编码，10个由核DNA编码。在这项研究中，采用了RNA干扰方法，从而产生了小鼠A9细胞衍生物，其该复合体的核编码亚基IV（COX IV）表达受到抑制。该亚基的量减少至正常水平的86%至94%。对几种COX IV表达受抑制的细胞系进行详细的生物合成和功能分析，结果显示细胞色素c氧化酶复合体组装缺失，相应地，细胞色素c氧化酶依赖性呼吸和总呼吸减少。此外，细胞中线粒体细胞色素c氧化酶功能失调导致线粒体膜电位受损、ATP水平降低以及在半乳糖培养基中无法生长。有趣的是，COX IV表达的抑制也使细胞对凋亡敏感。这些观察结果提供了证据，证明COX IV亚基对于功能性细胞色素c氧化酶复合体具有重要作用，同时也表明细胞色素c氧化酶对氧化磷酸化具有严格的调控。最后，我们的结果进一步揭示了细胞色素c氧化酶复合体功能失调所导致疾病的致病机制。
